Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9.

Effect of orthodontic treatment

Journal section: Clinical Dentistry doi:10.4317/medoral.15.e924

Publication Types: Research

Effect of orthodontic treatment on saliva, plaque and the levels of

Streptococcus mutans and Lactobacillus

Edith Lara-Carrillo 1, Norma-Margarita Montiel-Bastida 2, Leonor Sánchez-Pérez 3, Jorge Alanís-Tavira 4

1
DDS, MSD Doctoral student; Department of Orthodontics, Research Center, School of Dentistry, Universidad Autónoma del
Estado de México, Toluca, Estado de México, Mexico
2
DDS, PhD Professor and Head, Research Center, School of Dentistry, Universidad Autónoma del Estado de México, Toluca,
Estado de México, Mexico
3
DDS, MSD, PhD Professor, Health Attention Department, Universidad Autónoma Metropolitana Unidad Xochimilco, Mexico
City, Mexico
4
DDS, MSD, PhD Professor, Research Center, School of Dentistry, Universidad Autónoma del Estado de México, Toluca, Estado
de México, Mexico

Correspondence:
Facultad de Odontología de la UAEM
Centro de Investigación y Estudios Avanzados en Odontología
Paseo Tollocan esq. Jesús Carranza, Col. Universidad
C.P. 50130, Toluca, Estado de México. México
laracaedith@hotmail.com
Lara-Carrillo E, Montiel-Bastida NM, Sánchez-Pérez L, Alanís-Tavira J.
Effect of orthodontic treatment on saliva, plaque and the levels of Strep-
tococcus mutans and Lactobacillus. Med Oral Patol Oral Cir Bucal. 2010
Nov 1;15 (6):e924-9.
http://www.medicinaoral.com/medoralfree01/v15i6/medoralv15i6p924.pdf

Article Number: 3163 http://www.medicinaoral.com/

© Medicina Oral S. L. C.I.F. B 96689336 - pISSN 1698-4447 - eISSN: 1698-6946
eMail: medicina@medicinaoral.com
Received: 11/09/2009 Indexed in:
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Abstract
Objectives: The aim of this study was to identify changes in the oral environment with clinical, salivary and bacte-
rial risk markers after placement of fixed orthodontic appliances on permanent dentition.
Material and Methods: With ethical approval, we used different techniques to analyzed clinical, salivary and bac-
terial risk markers in 34 patients (mean age, 16.7 ± 5.2 years), 14 males and 20 females; before starting orthodontic
treatment and 1 month after. Clinical risk markers (decayed, missing, and filled surfaces [DMFS], O’Leary´s
plaque index, and plaque pH); salivary markers (unstimulated and stimulated saliva flow rate, buffer capacity, pH,
and occult blood in saliva) and bacterial counts (Streptococcus mutans and Lactobacillus). Data were analyzed by
paired t-test and χ2 test.
Results: This study showed that orthodontic appliances increased the stimulated salivary flow rate (p=0.0001),
buffer capacity (p=0.0359), salivary pH (p=0.0246) and occult blood in saliva (p=0.0305). Bacterial levels in-
creased slightly after 1 month of treatment, without statistical significance. Between genders, initially we ob-
served differences in: stimulated saliva (p=0.0019), buffer capacity (p=0.0381) and plaque pH (p=0.0430); after
treatment the unstimulated saliva (p=0.0026) showed differences.
Conclusions: Orthodontic treatment changes the oral environmental factors, promotes an increase in stimulated
flow rate, buffer capacity and salivary pH, which augment the anti caries activity of saliva. In contrast, increased
occult blood indicated more gingival inflammation, apparently because augmented the retentive plaque surfaces
and the difficult to maintain a good oral hygiene, rinsed the bleeding in saliva by periodontal damage.

Key words: Saliva, oral health, orthodontic treatment.

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Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9.
Introduction didn’t have any systemic diseases, use of antibiotics at
Malocclusions are the 3rd most common of oral health
problems, and are associated with a number of com-
plications (1). Orthodontic treatment often can correct
these complications or at least prevent them from pro-
gressing; but it also holds some potential for harm to
teeth and periodontal tissues. For example oral hygiene
may be difficult to maintain during treatment, which
may lead to plaque accumulation and gingival inflam-
mation (2). It has been shown that orthodontic treatment
induces changes in the oral environment, with an in-
crease in the bacteria’s concentration, and alterations in
buffer capacity, pH acidity and salivary flow rate (3);
however, little is known about periodontal inflamma-
tion that results in occult blood in saliva and the acidity
of dental plaque.
Considerable time and effort has been spent on develop-
ing tests to identify individuals at risk for developing
dental caries and periodontal damage. Diverse tests ex-
ist based predominantly on quantitative estimation of
Streptococcus mutans and Lactobacillus, and on deter-
mination of quantity and quality of saliva (4-6).
The caries activity test Cariostat® (Dentsply-Sankin KK,
Tokyo, Japan) developed by Shimono, was designed to
measure the decrease of pH caused by bacterial action
in the dental plaque. It has been reported positive corre-
lations between caries activity test score and the counts
of Streptococcus mutans and Lactobacillus (7).
Actually exist another colorimetric salivary test used as
indicator at inflammation which involves determining
occult blood derived from the gingival tissue for evalu-
ates periodontal disease in initial stages, called Sal-
ivaster® (Showa Yakuhin Kako Co. LTD, Tokyo, Japan)
(8,9).
It is important to identify the changes in the oral envi-
ronment in patients undergoing orthodontic treatment
with fixed appliances. In some cases involving long
treatment duration, the clinicians are committed to pre-
serving the oral health of the patient. Actually, there ex-
ist few clinical studies that show the physiologic and
biological characteristics of saliva and the correlation
with clinical specific entities.
Therefore, the aim of the present study was to identify
changes in the oral environment with clinical, salivary
and bacterial risk markers after placement of fixed or-
thodontic appliances on permanent dentition.
Material and Methods
Patients who required treatment with fixed orthodontic
appliances (straight wire technique) from the Orthodon-
tic Clinic at the Research Center and Advanced Studies
in Dentistry of the Autonomous University of the State
of Mexico (UAEM) were studied. Thirty-four patients:
14 males (mean age, 16.2 ± 3.4 years) and 20 females
(mean age, 17.2 ± 6.3 years) participated. These patients
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Effect of orthodontic treatment
least 15 days before initiating the study, active caries,
and on mixed dentition phase were excluded. This re-
search was approved by the Ethics Committee of the
University. All patients or their tutors consented to par-
ticipate after obtaining information about the study.
Samples were taken from each patient in two stages,
one before beginning the orthodontic treatment and
the other 1 month after placement of the appliances,
because this is the time for the appointment to change
the first arch wire. The orthodontics attachments were
placed in both arches.
The patients avoided eating or drinking and no tooth
brushing at least 2 hours before taking the samples in
both stages. All procedures were done by one calibrated
researcher, and commercial kits were used according to
the manufacturer’s instructions.
Intraexaminer reliability in using these kits and all
markers was tested prior to the study using a group of
orthodontic patients from one author´s (NMMB) clini-
cal practice. Dental and skeletal diagnoses were ob-
tained from each patient, defined by Angle’s molar class
and the subspinale-nasion-supramentale cephalometric
angle (ANB).
Recommendations to avoid retentive, cariogenic, or
hard foods during orthodontic treatment were provided
verbally. We recommended that patients brush their teeth
three times a day using the Bass modified technique
with toothpaste containing fluoride after placement of
appliances.
2.1 Clinical markers
• The decayed, missing and filling surfaces index
(DMFS) was recorded according to criteria of the World
Health Organization for permanent dentition.
• The supragingival plaque was disclosed with a chewable
tablet (Viarden®; Viarden, Mexico City, Mexico) and its
amount estimated using criteria according to O´Leary´s
plaque index (10,11).
Clinical markers only were measurement at the begin-
ning of the study.
2.2 Salivary markers
• Unstimulated saliva was the formation time (in
seconds) of small saliva drops in the inner mucous of
the lower lip (Saliva Check®, GC America Inc., Alsip,
IL, USA) (6).
• Stimulated saliva was the total stimulated saliva
obtained during 5 minutes by chewing an unflavored
piece of wax; the result was expressed in mL/min (6).
• Buffer capacity was determined by placing stimulated
saliva using a pipette, dispensed one drop onto each of
the 3 test pads in the reactive strip of the Saliva Check®
test. After 2 minutes, the final result was calculated by
adding the points according to the final colour of each
pad: very low = 0 to 5 points; low = 6 to 9 points; and
normal/high = 10 to 12 points (6).
Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9.
• To determine salivary pH, the reactive strip of the
Saliva Check® was submerged in stimulated saliva for
10 seconds, the color obtained was compared with the
chart: highly acidic = 5.0 to 5.8; moderately acidic = 6.0
to 6.6; and healthy saliva = 6.8 to 7.8 (6).
2.3 Bacterial markers
• To estimate the number of colony-forming units of
Streptococcus mutans counts (SM) per milliliter of sa-
liva (CFU/mL) was used Dentocult® SM (Orion Diag-
nostica, Espoo, Finland) according to Jensen and Brat-
thall (4). Counts were categorized as follows score: 0
= negative or <104 colony-forming units (CFU)/mL, 1
= 105 CFU/mL, 2 = 105 to 106 CFU/mL, and 3 = > 106
CFU/mL.
• The number of Lactobacillus was obtained by the use
of Dentocult® LB (Orion Diagnostica, Espoo, Finland),
described by Larmas (5,12). The growth densities were
categorized as follows: NC = no count, 0 = negative or
103 CFU/mL, 1 = 104 CFU/mL, 2 = 105 CFU/mL, and 3
= 106 CFU/mL.
2.4 Plaque pH
• The ability of dental plaque to produce acid (pH) was
determined by Cariostat® (Dentsply-Sankin KK, Tokyo,
Japan). Plaque was collected from buccal surfaces of first
upper molars, using a sterilized cotton swab supplied in
the kit, which was put into a test medium and incubated.
The test color change was compared with the pattern
provided by the manufacturer as follows: negative value
= pH 5.8-7.2; one positive value = pH 5.4 ± 0.3; two
positives value = pH 4.8 ± 0.3 and three positives value
= pH < 4.4 (7,13).
2.5 Occult blood in saliva
• To measure occult blood, the Salivaster® reactive strip
(Showa Yakuhin Kako Co. LTD, Tokyo, Japan) was
used. The procedure involves dipping the test paper in
stimulated saliva for 2-3 seconds and then judging by
comparing to the standard color change chart, divided
into 3 levels: without change = 0.0 mg of blood per dL
of saliva (no periodontal disease), light blue = 1.0 mg/dL
(incipient periodontal disease), and dark blue = 2.5 mg/
dL (periodontal disease present) (9).
Statistical analysis
Clinical data were only obtained before treatment
(DMFS and plaque index). For the other variables a
paired t-test or χ2 test were used to determine significant
differences between the first and second samples, and
between genders. The information was analyzed using
JMP 7.0 (SAS Institute Inc., Cary, NC, USA). The
statistical level of significance was set at 0.05.
Results
Thirty-four subjects participated in this study. Fifteen
patients had molar relation class I, fourteen had class II,
four had class III, and in one patient it was not possible
to determine the relationship due to absent first molars.
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Effect of orthodontic treatment
As for the skeletal diagnosis, fourteen were class
I, nineteen were class II, and one were class III. The
dental and skeletal diagnoses were not associated or
determined the behavior of any of the studied variables
(p>0.05) (data not shown).
Clinical markers
The mean DMFS index of the subjects was 6.47, although
it was greater in females (8.70) that in males (3.28), these
differences were not statistically significant (p=0.1352).
The mean O’Leary’s plaque index was 44.6 %, males
presented a slightly greater plaque percentage (50.84%)
than females (40.15%) (p=0.1809).
Salivary markers
We did not find statistically significant differences
in the unstimulated salivary production before and
1-month after the placement of orthodontic appliances,
nevertheless in the second stage there were differences
between gender in salivary rate (p=0.0026); specifically,
the unstimulated salivary production was lower in
women (Table 1).
The placement of orthodontic appliances promoted
a major stimulated salivary flow in the subjects, with
significant differences in the salivary production before
and after treatment (p=0.0001). The salivary stimulated
flow rate was greater in males at the beginning of the
treatment (p=0.0019; (Table 1).
The salivary buffer capacity showed differences after
placement of appliances (p=0.0359) and between
genders before treatment (p=0.0381), females showed
lower capacity (Table 1).
Significant differences were observed in the salivary pH
before and after treatment (p=0.0246) with an increase
of the pH value (Table 1).
Bacterial markers
We observed changes in the CFU of SM after placement
of appliances. Before treatment 14/34 subjects had high
values (>105); after one month of banding, 16/34 had
high values.
In the first sample 7/34 subjects had high levels (>105)
of Lactobacillus, in the second stage we found 20/34
subjects in these same level, although statistically
significant differences were not observed in this
distribution (p=0.6905; (Table 2).
Plaque pH
The acidity of the initial plaque no registered significant
modifications after placement of appliances (p=0.5467);
however, we found differences between genders in
the initial sample (p=0.0430); with negative values
predominating in females, and one positive values in
males. The second sample showed more subjects with
one positive values in both genders (Table 2).
Occult blood in saliva
Differences were observed in the gingival bleeding
before and after orthodontic treatment (p=0.0305), in
the second stage the bleeding in saliva increased. In the
Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9. Effect of orthodontic treatment

Table 1. Distribution of salivary markers by gender in the study (n = 34).

Stages
Marker Gender Before After pa
Salivary markers
Male 39.85 + 19.17 38.64 + 14.90
Unstimulated saliva (seconds) Female 45.65 + 19.03 57.35 + 17.43 0.4073
pb 0.3903 0.0026*
Male 1.72 + 0.65 1.76 + 0.80
Stimulated saliva (mL/min) Female 1.06 + 0.48 1.36 + 0.50 0.0001*
pb 0.0019* 0.0835
Male 8.78 + 2.19 8.71 + 2.16
Buffer capacity Female 6.90 + 2.69 7.70 + 1.59 0.0359*
pb 0.0381* 0.1247
Male 7.68 + 0.17 7.74 + 0.09
Salivary pH Female 7.53 + 0.38 7.73 + 0.09 0.0246*
pb 0.1672 0.7039
Data shown as mean + SD
pa value between before vs. after placement of appliances, based on paired t-test
pb value between genders, based on paired t-test
*p < 0.05

Table 2. Distribution of bacterial markers, plaque pH and occult blood in saliva by gender in the study (n = 34).

Stages

pa
Marker Gender Before After

Salivary markers

Male 39.85 + 19.17 38.64 + 14.90

Unstimulated saliva (seconds) Female 45.65 + 19.03 57.35 + 17.43 0.4073

pb 0.3903 0.0026*

Male 1.72 + 0.65 1.76 + 0.80

Stimulated saliva (mL/min) Female 1.06 + 0.48 1.36 + 0.50 0.0001*

pb 0.0019* 0.0835

Male 8.78 + 2.19 8.71 + 2.16

Buffer capacity Female 6.90 + 2.69 7.70 + 1.59 0.0359*

pb 0.0381* 0.1247

Male 7.68 + 0.17 7.74 + 0.09

Salivary pH Female 7.53 + 0.38 7.73 + 0.09 0.0246*

pb 0.1672 0.7039

NC = negative at culture
pa value between before vs. after placement of appliances,
based on X2 test
pb value between genders, based on X2 test
* p < 0.05

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Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9.
beginning of the study most of the subjects were in the
intermediate level with 1 mg/dL of occult blood in saliva
(incipient periodontal disease), however one month later
the periodontal disease present increased (Table 2).
Discussion
This study shows the changes in clinical, salivary and
bacterial markers that occur in the oral environment at
the beginning of orthodontic treatment with fixed appli-
ances on permanent dentition.
The following markers emerged as protective factors:
patients without active caries injuries increased sig-
nificantly stimulated salivary flow, buffer capacity, and
salivary pH, after placement orthodontic appliances.
In contrast, the following markers were negative risk
factors to the oral environment: slightly increase in the
infection levels of SM and Lactobacillus, and of occult
blood in saliva.
Oral environment has the capacity of adjustment to the
presence of a foreign body, increasing the salivary flow
which contributes to the autoclisis and modifying the
salivary composition to raise the pH and buffer capac-
ity, it prevents colonization by potentially pathogenic
microorganisms by denying them optimization of envi-
romental conditions.
In the present study, stimulated salivary flow was
greater in males, which is similar to international re-
ports (3,14,15). The differences have been attributed to
two theories: women present smaller salivary glands in
comparison with men and the female hormonal pattern
may contribute to diminished salivary secretion. The
mean salivary flow rate for both genders was found in
normal parameters: 1-3 mL/min in stimulated saliva and
from 0.25-0.35 mL/min in unstimulated saliva (16). The
variability of salivary flow rate has been established by
other researchers (14-17). Mechanical or chemical stim-
ulus is associated with increased salivary secretion, our
results support a direct and prolonged stimulatory effect
after one month of treatment with fixed orthodontic ap-
pliances on salivary flow.
Salivary buffer capacity presented a significant increase
in females after orthodontic therapy, situation that has
been reported by Chang et al. (3). Males showed higher
buffer capacity than females, this difference between
genders have been demonstrated previously (16). This
salivary function avoids the settling of pathogenic micro-
organisms in mouth and neutralizes the acids produced
by acidogenic bacterias, preventing enamel desminerali-
zation. This is one of the best risk indicators, because it
reveals the host response (5). Patients with a high buffer
capacity are often quite resistant to the caries process, be-
cause high host response can even compensate for active
caries habits. If orthodontic patient at high caries risk can
be identified at the beginning of treatment, preventive
measures can be implemented with maximal effect.
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Effect of orthodontic treatment
Salivary pH demonstrated a significant increase in the
1st month of treatment, as opposed to other studies in
which it has been established that pH suffers alterations
after 3 months of orthodontic treatment (3). Orthodontic
appliances increase the retentive plaque surfaces, caus-
ing elevated acid levels of concentration of hydrogen
ions in oral environment, then the pH decrease. How-
ever, we observed higher flows of stimulated salivary
secretion, which raise the concentration of bicarbonate
ions, then the pH also rises and the buffering power of
the saliva increases dramatically; demonstrating the
physiologic response to maintain the oral health in ad-
verse situations.
In this study, clinical markers showed that males had
lower DMFS index and higher O’Leary’s plaque index
than females. Males showed more acid plaque pH at
the beginning of treatment, nevertheless an increase in
acidity was demonstrated 1 month later in females. The
plaque pH had significant differences between genders
when the patients already were in orthodontic treat-
ment. The orthodontic appliances protected the plaque
from the tooth brushing action, the mastication, and the
salivary fluid. Accumulating more on the cervical re-
gion of the brackets or below the arches wire, which is
the area where a major demineralization can be found
(18). The plaque control is one of the most important
factors that limit the implantation and settling of causal
microorganisms from caries and periodontal disease.
In relation to bacterial counts, the present study showed
a slightly increase of colony formation, after placement
of appliances, as opposed to report previously (3).
However, the increase of retentive plaque surfaces might
cause a higher risk for the patient during the treatment,
because exist some stages in which more orthodontic
appliances can include.
Before initiating treatment, the majority of patients
showed incipient periodontal disease, possibly caused
by crowding, which is the principal motivation for the
orthodontic treatment. Likewise, a significant increase of
periodontal disease was observed 1 month after initiating
orthodontic therapy (2.5 mg/dL of saliva). According to
reported in the literature, the adjacent connective tissue
inflammation has been considered a consequence of the
use of orthodontic bands, determining that the condition
of the gum deteriorates during the treatment with fixed
appliances, even in patients with good oral hygiene (1).
Several preventive strategies relating to changes in the
oral environment in orthodontic patients were estab-
lished. Checking the quality, pH and buffer capacity of
saliva can be valuable as part of an overall clinical as-
sessment, thus also monitoring bacterial counts, plaque
and periodontal inflammation.
Med Oral Patol Oral Cir Bucal. 2010 Nov 1;15 (6):e924-9. Effect of orthodontic treatment

Conclusions 16. Elishoov H, Wolff A, Kravel LS, Shiperman A, Gorsky M.

Association between season and temperature and unstimulated
Orthodontic treatment changes the oral environmental
parotid and submandibular/sublingual secretion rates. Arch Oral
factors: promotes a major salivary stimulated flow and Biol. 2008;53:75-8.
increases its buffer capacity and salivary pH, which in- 17. Torres SR, Nucci M, Milanos E, Pereira RP, Massaud A, Munhoz
crease the anticaries activity of saliva. T. Variations of salivary flow rates in Brazilian school children. Braz
Oral Res. 2006;20:8-12.
Plaque pH did not demonstrate significant changes be-
18. Gwinnett AJ, Ceen RF. Plaque distribution on bonded brackets: a
fore and 1-month into orthodontic treatment. The bac- scanning microscope study. Am J Orthod. 1979;75:667-77.
terial levels did not increase significantly in the first
month of the orthodontic treatment but, the increase of Acknowledgments
retentive plaque surfaces and the difficult to remove it, We thank to the National Council of Science and Technology of Mex-
rinsed the bleeding in saliva by periodontal injury. ico (CONACYT) for their support.
It is necessary to maintain the balance between the pro-
tective factors and the caries risk factors during the or-
thodontic treatment with a rigorous home care program
toward correct oral hygiene procedures necessary to
control plaque accumulation for caries and periodontal
disease prevention.

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